Hypertension (high blood pressure) has been reported to be one of the most important risk factors associated with heart attack in industrialized countries. Hypertension is frequently treated with drugs that strongly inhibit the angiotensin-converting enzyme (ACE). The prevention of high blood pressure in the early stage of the development of the disease, can be an alternative, to the treatment of hypertension with drugs. A large number of food-derived bioactive compounds are currently considered as beneficial for general well being or as health promoting.
In the regulation of blood pressure, angiotensin I-converting enzyme (ACE) plays an important role. ACE acts to increase the blood pressure. In the renin-angiotensin system, ACE converts angiotensin-I to angiotensin-II by hydrolysing His-Leu from its C-terminal. Angiotensin II exhibits a strong vasoconstricting action. Additionally, in the kinin kallikrein system, ACE deactivates bradykinin, which aids vasodilation. ACE inhibitors are therefore useful in reducing blood pressure. Currently several ACE inhibitors already exist. The first reported ACE inhibitors were naturally occurring peptides found in snake venom. Since then, many other ACE inhibitors have also been discovered.
It is known that milk fermented by lactic acid bacteria (LAB) may produce anti-hypertensive effects due to the liberation of peptides from casein in the milk by the proteolytic activity of the lactic acid bacteria. The peptides act as ACE inhibitors.
The article of the company Calpis Food Industry [Yamamoto et al (1994, J. Dairy Sci., 77: 917-922)] discloses that milk fermentation by a Lactobacillus helveticus CP790 strain produced anti-hypertensive effects due to the liberation of peptides from casein in the milk by the proteolytic activity of L. helveticus. The peptides act as ACE inhibitors. The anti-hypertensive activity of these peptides was tested on spontaneously hypertensive rats. Milk fermentation by an isogenic mutant of Lb. helveticus that does not have proteolytic activity does not show any anti-hypertensive effects.
The article [Gobbetti M. et al (2000, Appl Environ Microbiol, 66 (9), 3898-3904.], describes fermented milk containing ACE-inhibitory peptides that were produced by using either Lactococcus lactis subspecies cremoris FT4 or Lactobacillus delbrueckii subspecies bulgaricus SS1 to ferment the milk.
EP821968 (Calpis Food Industry) describes fermented milk containing ACE-inhibitory peptides that were produced by using a Lactobacillus helveticus strain with deposit accession number FERM BP-4835.
EP1016709 (Calpis Food Industry) describes fermented milk containing ACE-inhibitory peptides that were produced by using a Lactobacillus helveticus strain with deposit accession number FERM BP-6060.
WO01/32836 (Valio Ltd) describes fermented milk containing ACE-inhibitory peptides that were produced by using a Lactobacillus helveticus strain with deposit accession number DSM 13137.
Because lactic acid bacteria (LAB) are auxotrophic for a number of amino acids, LAB depend upon a complex proteolytic system to obtain essential amino acids from casein during growth in milk. The hydrolysis of casein into amino acids for use by LAB is initiated by cell wall proteinases that hydrolyse casein into oligopeptides. Oligopeptides are then transported into the bacterial cell via an oligopeptides transport system. Once the oligopeptides are inside the cell, intracellular peptidases hydrolyze them to free amino acids.
The article of University of Wisconsin and Utah State University [Pederson et al (1999, J. of Bacteriology, 181: 4592-4597] describes the DNA and amino acid sequence of a prtH 204 kDa cell wall proteinase from the Lactobacillus helveticus CNRZ32 strain. This article does not describe nor suggest using of the strain to make peptides with anti-hypertensive properties.
The article of the company Calpis Food Industry [Yamamoto et al (2000), Biosci. Biotechnol. Biochem., 64(6): 1217-1222] describes the DNA and amino acid sequence of a prtY 45 kDa cell wall proteinase from the Lactobacillus helveticus CP790 strain. The CP790 strain does not comprises the prth 204 kDa cell wall proteinase [see “Discussion” section of Yamamoto et al (2000)]. The CP790 strain is used in a commercial product of Calpis Food Industry as the Materials and Methods sections reads, “CP790, was isolated from the starter culture of Calpis, a Japanese fermented milk product”.